The objectives of this project are to improve definition of growth control mechanisms in nontransformed cells, alterations that occur in cells neoplastically transformed by chemicals, and the molecular basis for these control mechanisms. Conditions for the serum-free culture, growth arrest, and stimulation of mouse embryo-derived AKR-2B cells and their chemically transformed derivatives (AKR-MCA) have been developed. The growth factor requirements for the AKR-2B cells have been established. It has been shown that transforming growth factor, type beta (TGF-beta), that stimulates growth of large colonies in soft agar is also a mitogen for these cells in monolayer in contrast to previous reports. TGF-beta induces DNA synthesis with a longer lag phase between stimulation and the onset of DNA synthesis than with other growth factors so that previous assays were performed too early. TGF-beta is not only a unique mitogen in that it stimulates a later S phase, but also it suppresses the effect of other growth factors before exerting its own effect on proliferation. Two complementary DNA libraries have been constructed in lambda GT-11 phage, one from mRNA from cells stimulated with epidermal growth factor (EGF) and another from cells stimulated with TGF-beta. These are presently being screened for sequences specifically induced by EGF and TGF-beta. Several clones induced by EGF (that are not VL30-related) have been selected and are undergoing secondary and tertiary screening. Other studies on gene expression involve the use of known cloned genes to examine growth factor induction. TGF-beta has been shown to induce VL30, actin, and the JE and KC clones isolated by Stiles and co-workers as platelet-derived growth factor inducible genes. Analysis of cytosolic proteins by gel electrophoresis following [unreadable]35[unreadable]S-methionine labeling has shown two proteins (one acidic protein of approximately 38 kilodaltons, another more basic protein of about 50 kilodaltons) that are specifically induced by TGF-beta but not by EGF. (J)